B lymphocytes are subject to many checkpoints in self-tolerance enforced by at least three mechanisms that include anergy, receptor editing and clonal deletion. Nevertheless, in certain autoimmune diseases such as systemic lupus erythematosus (SLE), they somehow manage to evade all of these checkpoints, multiply by clonal expansion and produce high-affinity autoantibodies that are products of T cell-dependent immunity. Recent experiments from this laboratory provide important insight regarding this paradox. Our data conclusively show for the first time, in a spontaneous mouse model of SLE, that lupus anti-nuclear B cells are frequently created de novo from normal nonautoreactive precursors via the process of somatic hypermutation (SHM). As such, they are able to traverse all of the early preimmune checkpoints in tolerance because they are not autoreactive until they acquire somatic mutations in the periphery, presumably during germinal center (GC) reactions. This finding underscores the importance of self-tolerance checkpoints that occur at late stages of B cell differentiation during humoral immune responses. Therefore, to understand how mutant autoimmune B cells are normally regulated under physiological circumstances, and how they escape self-tolerance under disease circumstances, we propose to generate a novel Ig gene-targeted model that will allow us to analyze self-tolerance mechanisms that operate during immunity. This is an important endeavor because all current Ig transgene or gene-targeted models of anti-nuclear B cells employ cells that express autoreactive receptors from the moment they arise during lymphopoiesis in the bone marrow. As such, the B cells are tolerant to varying degrees prior to antigen encounter. The model we propose will make use of CRE recombinase-mediated deletion to convert a nonautoreactive B cell into an autoreactive B cell as it participates in an immune response, thus simulating an event that occurs naturally via SHM. The autoreactive B cells thus generated will become labeled with GFP. In addition, they will express a receptor originally derived from an actual autoimmune B cell that arose spontaneously in a mouse with SLE. The control of B cell specificity afforded by this model will enable us to define self-tolerance checkpoints at the GC and post-GC stages of development under physiological conditions, and conversely mechanisms of escape in autoimmunity, at an unprecedented level of resolution. This model is essential if we are to understand how autoreactive B cells evade the final checkpoints in self-tolerance so that rational therapies can ultimately be designed to silence them physically or functionally. PUBLIC HEALTH RELEVANCE: B lymphocytes that produce antibodies directed against self-tissues are major participants in systemic autoimmune diseases such as systemic lupus erythematosus (SLE). This project aims to develop a unique state-of-the-art-model to understand how these potentially dangerous cells are normally contained or eliminated under physiological circumstances and conversely how they sometimes evade control mechanisms to participate in autoimmune disease. The information obtained from this model will provide a basis for the rational design of therapies to control or prevent autoimmune diseases in which B cells play a significant role.